Differential effects of vanadate on UDP- N-acetylglucosaminyl transferase activity derived from cytosol and nucleosol

Abstract

UDP- N-acetylglucosaminyl transferase (OGT) is a key enzyme of a novel signal transduction pathway that regulates protein function through O-linked glycosylation. In the current study, we found that sodium vanadate potently inhibits OGT activity in brain cytosol (IC 50= 55 μM) and nucleosol (IC 50= 150 μM), but fails to alter activity of a related enzyme (UDP-galactosyltransferase). Vanadate also inhibits OGT activity in cytosol (IC 50 of 2.3 μM) and nucleosol (IC 50 of 130) derived from a stable HeLa cell line that overexpresses OGT. When HeLa cytosol was immunopurified to separate OGT from other cellular proteins, vanadate still inhibited OGT activity (IC 50= 2 μM). We conclude that OGT derived from cytosol exhibits greater vanadate sensitivity than nucleosol OGT and that a large difference exists (25-fold) in vanadate sensitivity when comparing OGT activity in different cell types (IC 50 of 55 μM for brain cytosol vs. 2.3 μM for HeLa cytosol). Understanding the mechanism(s) by which a tyrosine phosphatase inhibitor differentially reduces OGT activity should lead to new insights into OGT function and regulation.